ABSTRACT Psychiatric diseases and addictive behaviors are complex brain disorders associated with maladaptive plasticity of the brain circuitry. The lack of adequate platforms to rapidly screen against novel targets in physiological environments has significantly hampered probe discovery initiatives that could inform the circuitry alterations leading to mental disorders and addiction and enable the design of future therapeutics. Protein- protein interactions (PPI) within ion channel complexes fine-tune neuronal excitability and are emerging as links to the biology of psychiatric disorders. Their highly specific and flexible interfaces could make protein- channel interactions ideal targets for probe development. Such molecular probes would provide the neuropharmacology community with optimal research tools to parse out brain disease complexities and ultimately enable future drug design. We have identified the PPI between the voltage-gated Na+ (Nav) Nav1.6 channel and its accessory regulator protein, fibroblast growth factor 14 (FGF14) as a novel, functionally relevant regulator of neuronal excitability in brain areas such as the cortico-mesolimbic circuit, which is associated with disorders of the affective and cognitive domains. Through a successful bioluminescence-based high-throughput screen (HTS) and subsequent in vivo studies in the nucleus accumbens (NAc), we discovered that the FGF14:Nav1.6 channel complex is part of the glycogen synthase kinase 3 (GSK3) pathway, a signaling cascade found aberrant in bipolar disorder, depression, anxiety and addiction. To explore the druggability of the FGF14:Nav1.6 complex, we employed a minimal functional domain (MFD) approach to design a peptide-derivative mapped to the PPI interface and showed it has in vitro-to-ex vivo activity in the NAc circuit. These discoveries have prompted us to develop a new pipeline to identify chemical probes against the FGF14:Nav1.6 complex to interrogate its function in the cortico-mesolimbic circuit. To advance a probe discovery campaign against this new target, we have designed an integrated multi-modal screening platform, based on the latest MFD principles of pharmacology that includes a newly designed and validated bioluminescence primary screening assay to reconstitute the FGF14:Nav1.6 C-tail complex in cells. This pipeline also includes the necessary counter, toxicity, and cell-free orthogonal assays (Aim 1), automated patch-clamp electrophysiology as a functional screen in combination with structure-activity relationship efforts and in silico analysis (Aim 2), and ex vivo validation of selected probes in the NAc circuitry (Aim 3). The proposed pipeline introduces a new, rapid and integrated platform that will accelerate the discovery of novel chemical probes for neuronal excitability, providing the foundation for pre-therapeutic development of a new class of PPI-based leads for a broad spectrum of psychiatric disorders.